Study On Injectable Tissue-engineered Bone With Calcium Alginate Gels As Carriers

Bone defect caused by tumour, inflammation, trauma, congenital and growth malformation is one of the common diseases in the clinic. The traditional repairing way has deficiency to some extent. The advent of tissue engineering lights a new approach for repairing and reconstruction of bone defect. The basic principle of tissue engineering is applied to fabricate injectable tissue-engineered bone. Tissue-engineered bone with injectable excellular scaffold material has the advantages of slight damage and easy handling, which is becoming one of the focuses in the current tissue engineering research. The research of injectable tissue-engineered bone has been conducting mainly in the following three aspects: 1. Researchs on conditions and environment of seed cells cultivated in vitro. 2. Researchs on developing and improving the property of three-dimension biological scaffold materials. 3. Research on construction of injectable tissue-engineered bone. This experiment focuses on the above three aspects. A series of research about calcium alginate gels as carriers has been carried on, which provides, as we hope, theoretical basis for the clinic application of injectable tissue-engineered bone as soon as possible.Part I Observation of proliferation, differentiation, and bioaction of dog marrow stromal cells1. Cultivation and identification of osteoblasts from bone marrow stromal cells of dog in vitro Objective: To observe the osteoblastic differentiation and osteogenetic phenotype from bone marrow stromal cells of dog in vitro. Methods: Bone marrow stromal cells obtained from dog ilium through puncture were cultured in DMEM medium (containing 100ml/L fetal bovine serum) and mineralizational medium (containing Dex, Vit C and Na β -glycerophosphate). The osteoblastic differentiation, proliferation and osteogenetic phenotype were observed. Result: Bone marrow stromal cells cultured in mineralizational medium were more healthy than those cultured in DMEM medium. Osteogenetic phenotype was observed after the bone marrow stromal cells were cultured in mineralizational medium, but the growth rate of the cells slowed down. Conclusion: Bonemarrow stromal cells obtained through puncture could be differentiated into osteoblasts by culturing in mineralizational medium.2. Bioaction of bone marrow stromal cells with calcium alginate gels in vitro Objective: To observe the bioaction of bone marrow stromal cells in calcium alginate gel. Method: Dog bone marrow stromal osteoblasts which had been differentiated and proliferated in vitro were mixed with sodium alginate solution to generate an osteoblasts/calcium alginate gel composite. After 1 to 5 days culture, histomorphological analysis, electron microscopy and immunohistochemistry were performed to evaluate the phenotypic changes of osteoblasts as well as their living condition in calcium alginate gel. Result: Bone marrow stromal osteoblasts embedded in three-dimension calcium alginate gel were living in a floating condition. There were a few extracellular matrices were observed by SEM after 5 days. Osteoblasts' phenotype was maintained. Conclusion: Phenotype of osteoblasts was maintained in calcium alginate gel and a few matrices were secreted, which indicates that calcium alginate gel has potential for tissue engineering bone. Part II Study on mechanical and chemical properties of calcium alginate gels1. Determination of mechanical and chemical properties of calcium alginate gelsObjetive: To apply calcium alginate gels in injectable tissue-engineered bone reasonably, the pH of alginate solution and mechanical properties of alginate gels were measured. Method: The 30g/L alginate solution was made with alginate and its pH was measured with pHS-2 acidity meter. The calcium alginate gels with defined dimensions were achieved by adding CaSO4 into alginate solution. Then the compressive strength, tensile strength, and flexural strength were measured on omnipotence material experimental machine. Result: The pH of alginate solution is 7.332. The compressive streng...